A Hybrid Viral/Nonviral Vector for CFTR Delivery to CF Pig Airways

用于 CFTR 递送至 CF 猪气道的混合病毒/非病毒载体

• 批准号: 9923461
• 负责人: PATRICK L SINN
• 金额: $ 57.52万
• 依托单位: UNIVERSITY OF IOWA
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-01 至 2022-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9923461
• 关键词: Address; Adenovirus Vector; Animal Model; Anions; Basal Cell; Biliary cirrhosis; Categories; Cell surface; Cells; Characteristics; Clinical; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; DNA; DNA Transposons; Data; Defect; Disease; Disease Outcome; Disease Progression; Dose; Electrolytes; Emerging Technologies; Engineering; Epithelial Cells; Exhibits; Exocrine pancreatic insufficiency; Failure; Family suidae; Gene Delivery; Gene Expression; Gene Transfer; Gene therapy trial; Genes; Genetic Diseases; Genome; Genomics; Goals; Guide RNA; Host Defense; Human; Hybrids; Ileus; In Vitro; Infection; Inflammation; Intestines; Ion Transport; Lentivirus Vector; Life; Lobe; Lung; Lung diseases; Measures; Meconium; Mendelian disorder; Methods; Modeling; Mucous body substance; Mus; Neonatal; Newborn Infant; Non-Viral Vector; Nose; Outcome; Phenotype; Population; Pulmonary Cystic Fibrosis; Quality of life; Reagent; Reporter Genes; Reporting; Research; Research Personnel; Structure of respiratory epithelium; Surface; System; Testing; Therapeutic; Transgenes; Transposase; Treatment Efficacy; Viral; Viral Vector; adeno-associated viral vector; aerosolized; airway epithelium; airway obstruction; base; cell type; cellular transduction; cystic fibrosis mouse; disease phenotype; disease-causing mutation; disorder prevention; efficacy testing; experimental study; falls; gene replacement; gene therapy; genetic approach; gutless adenoviral vector; improved; in vivo; innovation; knockout gene; novel; novel strategies; overexpression; preclinical study; prevent; progenitor; promoter; stem cells; success; therapeutic evaluation; therapeutic gene; tool; uptake; vector

Project Summary/Abstract Gene therapy was first proposed as a viable therapeutic strategy for genetic disease nearly 30 years ago. There are many notable recent successful gene therapy trials for multiple diverse diseases; however, gene therapy for cystic fibrosis (CF) has presented many unexpected roadblocks. For example, 1) airway cells have evolved many barriers to prevent uptake of foreign DNA, 2) mouse models of CF generally lack lung disease, and 3) each of the standard gene delivery vehicles have pros and cons. We have a demonstrated track record of utilizing many categories of viral and non-viral based reagents for gene delivery to the airways. Viral based vectors are the most efficient way to deliver genes to airway cells. Our long-term goal is to engineer a reagent for gene delivery to cells that is an effective therapeutic for CF lung disease, yet there is a critical need for improved gene delivery tools. Here, we develop a novel reagent. We propose to demonstrate that our engineered vector can integrate into the genome of progenitor cells in the airways and express the transgene in the appropriate surface cells. To this end, we have combined the efficiency of an adenoviral-based vector with the persistent expression of a DNA transposon-based non-viral vector. In addition, we test this `hybrid' vector in a relevant large animal model of CF. Here we demonstrate that this vector will: 1) deliver CFTR to CF cells and correct the anion transporter defect in vitro using a novel strategy to focus cellular integration and expression, 2) verify that the necessary levels of gene transfer are achievable in pigs airways in vivo, and 3) correct the anion channel defect and disease progression in the airways of a pig CF model. We investigate a gene delivery tool that can be used to address questions that no other reagent can address. There are >2000 known disease causing mutations in CFTR. Our goal is to provide a life-long gene replacement strategy that would be efficacious regardless of the disease causing mutation. This proposed research is highly innovative. Combining the emerging technologies of DNA transposon-based vectors with well-studied adenoviral delivery presents an exciting new opportunity to improve the utility of delivery vehicles for therapeutic genes such as CFTR. The reagents, methods, and data generated by these experiments will provide guidance for gene therapies for other monogenic disorders, thereby significantly advancing the gene therapy field.

项目摘要/摘要 大约30年前，首先提出了基因疗法作为遗传疾病的可行治疗策略。 最近有许多针对多种多种疾病的成功基因疗法试验。但是，基因 囊性纤维化治疗（CF）提出了许多意外的障碍。例如，1）气道电池具有 进化了许多防止外国DNA摄取的障碍，2）CF小鼠模型通常缺乏肺部疾病， 3）每个标准基因输送车辆都有利弊。我们有一个示威的记录 利用许多类别的病毒和非病毒试剂将基因递送到气道。基于病毒 向量是将基因输送到气道细胞的最有效方法。我们的长期目标是设计试剂 对于CF肺部疾病的有效治疗的细胞基因递送，但至关重要 改进的基因输送工具。在这里，我们开发了一种新型试剂。我们建议证明我们的 工程矢量可以集成到气道中的祖细胞的基因组中，并表达转基因 在适当的表面细胞中。为此，我们结合了基于腺病毒的载体的效率 基于DNA转座子的非病毒载体的持续表达。此外，我们测试了此“混合动力” 相关的大型动物模型中的向量。在这里，我们证明该向量将：1）将CFTR传递到CF 细胞并使用新型策略在体外纠正阴离子转运蛋白缺陷，以聚焦细胞整合和 表达，2）验证在体内猪气道中可以实现必要的基因转移水平，而3） 纠正PIG CF模型气道中的阴离子通道缺陷和疾病进展。我们调查一个 可用于解决其他试剂无法解决问题的基因输送工具。有> 2000 已知疾病导致CFTR突变。我们的目标是提供终身基因替代策略 不管导致突变的疾病如何，都将是有效的。这项拟议的研究具有很高的创新性。 将基于DNA转座子的载体的新兴技术与研究良好的腺病毒递送结合 提供了一个令人兴奋的新机会，以改善用于治疗基因的递送工具的实用性，例如 CFTR。这些实验生成的试剂，方法和数据将为基因提供指导 其他单基因疾病的疗法，从而显着推进基因治疗领域。